Mitochondria are essential, multi-functional organelles that control metabolism, but are also targets of environmental stress and involved in human disease pathology. Mitochondria produce reactive oxygen species (ROS) that damage macromolecules, promote oxidative stress, and initiate cell death that is exacerbated by many environmental toxins. Mitochondrial ROS also participate in signal transduction mechanisms that can have both detrimental and beneficial outcomes, depending on the exact physiological or environmental context. However, the mechanisms of mitochondrial ROS sensing and the specific stress-response pathways that determine these differential outcomes are understudied and far from understood. A major limitation in understanding mitochondrial pathology, and how environmental toxins promote or exacerbate it, is the complex tissue-specificity involved in mitochondrial function and stress responses. There is also a dearth of animal models in which to generate and monitor acute and chronic mitochondrial stress and toxicology in a tissue-specific manner or at specific times in development. The major goal of the R21 portion of this proposal is to develop novel mouse models of mitochondrial stress via controlled AND reversible knock-down of the mitochondrial superoxide dismutase gene (SOD2), which will increase the levels of mitochondrial ROS, and hence model environmental toxin exposures and allow mitochondria-to-nucleus signaling pathway signatures to be identified systematically. Using these signatures, we will then determine if environmental tobacco smoke (ETS) exposure in mice evokes a mitochondrial ROS response as part of its toxic mechanism. In the R33 portion of this proposal, cutting-edge mitochondrial antioxidants will be used to more precisely define the mitochondrial ROS stress signatures obtained in the R21 Phase and to prevent the deleterious tissue effects of ETS and two of its highly toxic constituents, the polycyclic aromatic hydrocarbon B[a]P and the heavy metal Cd. Finally, these new mouse models will be used to test the concept that mitochondrial ROS produced during development result in adaptive signaling responses that determine the nature or degree of resistance to subsequent toxin exposures in adults. The results of this study will greatly expand knowledge of the role of mitochondria and ROS signaling in environmental stress-induced toxicity and the complex tissue-specific pathology involved, and will inform future studies aimed at monitoring, diagnosing, and perhaps counteracting these.